Adipocyte Fatty Acid-Binding Protein 4 Suppresses Contraction of Mouse Ventricular Myocytes via a Calcium-Independent Pathway

Cuihua Wang¹Daofeng You¹Xuan Jiang¹Shanshan Han¹Fenghong Liu¹Wei Wang²Mingqi Zheng^1*

• ¹The First Hospital of Hebei Medical University, Shijiazhuang, China
• ²Jiangxi Provincial People's Hospital, Nanchang, China

Objective: Adipocyte Fatty Acid-Binding Protein 4 (FABP4) exerts a direct negative inotropic effect on cardiac muscle, but the underlying cellular mechanisms remain elusive. This study aimed to dissect the specific effects of FABP4 on the contractility and calcium (Ca²⁺) homeostasis of isolated mouse ventricular myocytes and to characterize the functional role and critical residues of its N-terminal domain. Methods: Contractility and intracellular Ca²⁺ transients were simultaneously measured in isolated adult mouse ventricular myocytes using an IonOptix system following acute application of recombinant human FABP4 or its synthetic N-terminal peptide (FABP4ₐₐ₁₋₂₀). L-type Ca²⁺ current was assessed via the whole-cell patch-clamp technique. Dose-response curves were analyzed using non-linear regression, and site-directed mutagenesis (E15K) was performed to evaluate the functional importance of a key amino acid residue. Results: FABP4 inhibited myocyte contraction in a biphasic, dose-dependent manner, with a high-affinity (EC₅₀ = 0.010 pM) and a low-affinity (EC₅₀ = 0.120 nM) component. This inhibition was largely independent of Ca²⁺ handling, as Ca²⁺ transient amplitude was only weakly attenuated at higher concentrations (EC₅₀ = 0.412 nM), and L-type Ca²⁺ current was unaffected. In stark contrast, the FABP4ₐₐ₁₋₂₀ peptide also inhibited contraction (EC₅₀ = 0.110 nM) but did so via a Ca²⁺-dependent pathway, robustly suppressing Ca²⁺ transients. Mutation of glutamic acid at position 15 (E15K) significantly attenuated the peptide's inhibitory activity. Conclusion: Full-length FABP4 suppresses cardiomyocyte contractility primarily through a Ca²⁺-independent pathway, likely by reducing myofilament Ca²⁺ sensitivity. Conversely, its isolated N-terminal domain operates via a distinct, Ca²⁺-dependent mechanism. These findings reveal a complex dual-pathway regulation of cardiac function by FABP4 and identify its N-terminal region as a potential therapeutic target for mitigating obesity-related cardiac dysfunction.